Definition of amino acid residues on the epitope responsible for recognition by influenza A virus H1-specific, H2-specific, and H1- and H2-cross-reactive murine cytotoxic T-lymphocyte clones

Abstract

We defined the epitopes recognized by three influenza A virus-specific, H-2Kd-restricted CD8(+) cytotoxic T-lymphocyte (CTL) clones: H1-specific clone A-12, H2-specific clone F-4, and H1- and H2-cross-reactive clone B7-B7. The A-12 and B7-B7 clones recognized the same peptide, which comprises amino acids 533 to 541 (IYSTVASSL) of A/PR/8 hemagglutinin (HA). The F-4 and B7-B7 clones both recognized the peptide which comprise amino acids 529 to 537 (IYATVAGSL) of A/Jap HA. Amino acids 533 to 541 of A/PR/8 HA are compatible with amino acids 529 to 537 of A/Jap HA. Amino acid S at positions 3 and 7 was responsible for recognition by H1-specific clone A-12, while amino acid G at position 7 was responsible for recognition by H2-specific clone F-4. Two conserved amino acids, T at position 4 and A at position 6, were responsible for recognition by H1-, and H2-cross-reactive clone B7-B7. These results indicate that a single nine-amino-acid region is recognized by HA-specific CTL clones of three different subtype specificities and that the amino acids responsible for the recognition by the CTL clones are different.

FIG. 1

P815 cells were pulsed with each of peptides f1 to f8 at 10 μg/ml and used as target cells for H1-specific CTL clone A-12, H2-specific CTL clone F-4, and the H1- and H2-cross-reactive CTL clone B7-B7 in 4-h CTL assays. The effector-to-target cell ratios were 8:1 for the A-12 clone, 6:1 for the F-4 clone, and 7:1 for the B7-B7 clone.

FIG. 2

P815 cells were pulsed with each of peptides g1 to g8 and h1 to h3 at various concentrations and used as target cells for H1- and H2-cross-reactive CTL clone B7-B7 in a 4-h CTL assay. The effector-to-target cell ratio was 7:1 when peptides g1 to g8 were used and 4:1 when peptides h1 to h3 were used.